Profilins are small proteins that form complexes with G‐actin and phosphoinositides and are therefore considered to link the microfilament system to signal transduction pathways. In addition, they bind to poly‐L‐proline, but the biological significance of this interaction is not yet known. The recent molecular cloning of the vasodilator‐stimulated phosphoprotein (VASP), an established in vivo substrate of cAMP‐ and cGMP‐dependent protein kinases, revealed the presence of a proline‐rich domain which prompted us to investigate a possible interaction with profilins. VASP is a microfilament and focal adhesion associated protein which is also concentrated in highly dynamic regions of the cell cortex. Here, we demonstrate that VASP is a natural proline‐rich profilin ligand. Human platelet VASP bound directly to purified profilins from human platelets, calf thymus and birch pollen. Moreover, VASP and a novel protein were specifically extracted from total cell lysates by profilin affinity chromatography and subsequently eluted either with poly‐L‐proline or a peptide corresponding to a proline‐rich VASP motif. Finally, the subcellular distributions of VASP and profilin suggest that both proteins also interact within living cells. Our data support the hypothesis that profilin and VASP act in concert to convey signal transduction to actin filament formation.

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The focal adhesion protein VASP, a possible link between signal transduction pathways and the microfilament system, is phosphorylated by both cAMP-and cGMP-dependent protein kinases in vitro and in intact cells. Here, the analysis of VASP dephosphorylation by the serinelthreonine protein phosphatases (PP) PPI, PP2A, PP2B and PP2C in vitro is reported. The phosphatases differed in their selectivity with respect to the dephosphorylation of individual VASP phosphorylation sites. Incubation of human platelets with okadaic acid, a potent inhibitor of PPI and PP2A, caused the accumulation of phosphorylated VASP indicating that the phosphorylation status of VASP in intact cells is regulated to a major extent by serine! threonine protein phosphatases. Furthermore, the accumulation of phosphorylated cAMP-dependent protein kinase substrate(s) appears to account for inhibitory effects of okadaic acid on platelet function.